D ESCRIPTION (provided by applicant): CD40 signaling of dendritic cells (DCs) is so crucial that in its absence cell-mediated immunity is severely impaired. Contrary to early literature, emerging studies show that CD154-CD40 interactions alone are meager in inducing DC maturation. These recent studies show that the concomitant involvement of CD40 and other "co-inflammatory signals" are essential to induce maximal DC functions, a process referred to as DC "priming". We hypothesize that cross-talk amongst the signaling intermediaries of the CD40 cascade and these other inflammatory cascades (e.g. Toll-like receptor (TLR) cascade) may be responsible for priming and the optimizing of DC maturation. While CD40 agonists have been shown be marginally effective in vivo, the real potential of CD40 agonists have been underestimated because of their use in the absence of other in vivo priming signals. The specific aims are: Aim 1. Mapping the functional domains in the cytoplasmic tail of CD40 (CD40cyt) that are responsible for early signaling and alterations in DC function. We present a unique cohort of Tg mice (X-CD40) that express mutations in the CD40cyt which disrupt the interaction of the cytoplasmic domain with specific TRAFs. DCs from these X-CD40 mice will be studied to address the causal relationships between TRAF recruitment and the induction of in vitro /in vivo DC function. Secondly, the function of DCs isolated from genetically-deficient mice lacking TRAFs will be employed to confirm the functional contribution of these elements to CD40-induced DC biology. Aim 2. The basis for the cross-talk between the CD40 and TLR cascades in DO maturation. Studies are presented to illuminate the mechanisms underlying how maturation of DCs by concomitant engagement of TLR and CD40 synergize. Emphasis will be focused on the cross-talk between the CD40 and TLR cascades due to their profound synergy and shared use of a variety of signaling intermediaries. Aim 3. Synergy of the CD40 and TLR pathways in eliciting protective anti-tumor immune responses. The observations that CD40 agonists alone "short-circuit" both humoral and cellular immunity underscore the importance of DC "priming" in optimizing the use of CD40 as an adjuvant. We hypothesize that the combined use of TLR and CD40 agonists will be effective immunotherapeutic approaches to manage tumor models of established disease.